Anti-atherogenic functions of HDL include mediation of reverse cholesterol transport, and anti-oxidative and anti- inflammatory effects. Mounting evidence supports the concept that HDL function can be impaired or lost, and that dysfunctional HDL contributes to the development of atherosclerosis. Familial hypercholesterolemia (FH) is an autosomal dominant disorder associated with severely elevated LDL levels and increased risk of premature coronary artery disease (CAD). Both heterozygous and homozygous FH have low HDL, but this finding has not received much attention, and the increased CAD risk is attributed to the elevated LDL levels. We have recently discovered that the HDL from patients with FH is dramatically pro-inflammatory. A goal of this proposal (Specific Aim 1) is to examine the hypothesis that HDL of FH patients is dysfunctional (pro-inflammatory, pro- oxidant, and/or associated with reduced efflux capacity). Isoprostanes are formed from the non-enzymatic oxidation of arachidonic acid in phospholipids of lipoproteins and cell membranes. Plasma levels of F2- isoprostanes are established in vivo markers of lipid peroxidation and oxidant stress. Interestingly, the esterified isosprostanes, (EIs) in human plasma are mainly localized to HDL, with lesser amounts associated with LDL and VLDL. Several lines of evidence support a direct role for lipid peroxidation and/or F2-isoprostanes in atherosclerotic lesion development. At present it is unknown whether the accumulation of EIs in HDL affects its atheroprotective functions. Interestingly, we have found that plasma from subjects with FH undergoing LDL apheresis has twice the amount of EIs compared to normal plasma, and that after LDL apheresis EI levels decrease by 45%. Isolevuglandins (isoLGs) are a group of g-ketoaldehydes (isoketals) that are highly reactive mediators of oxidative damage formed in parallel with isoprostanes. We will investigate whether these bioactive lipids play direct mechanistic roles in the pro-inflammatory properties of HDL in FH patients. In Specific Aim 2, we will examine the hypothesis that accumulation of bioactive lipids (F2-isoprostanes and Isolevuglandins) in HDL results in loss of anti-inflammatory function. Isoprostanes have been shown to bind to eicosanoid receptors implicated in atherogenesis, including receptors for thromboxane (TP) and prostaglandin (PG)E2 (EP2). Therefore, we will examine the hypothesis that the TP and EP2 receptors contribute to the inflammatory effects of F2-isoprostane enriched HDL. Isoketals rapidly adduct to the e-amino groups of lysines on proteins, promoting protein cross-linking and dysfunction. Therefore, we will examine the hypothesis that isolevuglandins mediate oxidative damage of HDL associated proteins resulting in impaired function of HDL cholesterol. Our results may lead to the development of new markers of HDL function, provide new pharmacologic targets for improving HDL function, and identify HDL as a target of therapy in FH patients.